Method optimization in capillary zone electrophoretic analysis of hGH tryptic digest fragments.
The mobile phase composition was optimized for the separation of tryptic digest fragments of human growth hormone by capillary zone electrophoresis. The effect of pH (pH 2.4, 6.1, 8.1 and 10.4) was evaluated since pH determines the relative charge of species, the prime contributor to selectivity; pH 8.1 was selected for the optimization studies. Tricine (buffer), sodium chloride (ionic strength adjustor), and morpholine (mobile phase additive) concentrations were systematically varied a pH 8.1. All three exhibited major effects on the electroosmotic flow velocity and current, and minor effects on selectivity. Tricine was the most crucial for good resolution, although addition of morpholine helped to resolve closely eluting species. The optimum separation conditions were found to be pH 8.1 with 0.1 M tricine, 0.02 M morpholine and no salt.
Nielsen
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RG
Eli Lilly and Company, Lilly Research Laboratories, Lilly Corporate Center, Indianapolis, IN 46285.
Rickard
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EC
D004305:Dose-Response Relationship, Drug; D004586:Electrophoresis / Q000379:methods*; D013006:Growth Hormone / Q000302:isolation & purification* / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D009025:Morpholines / Q000494:pharmacology; D012965:Sodium Chloride / Q000494:pharmacology; D014357:Trypsin / Q000494:pharmacology*
D009025:Morpholines; D012965:Sodium Chloride; D013006:Growth Hormone; D014357:Trypsin